Molecular mechanism for sodium-dependent activation of G protein-gated K+ channels
Molecular mechanism for sodium-dependent activation of G protein-gated K+ channels
复制标题
DOI：
10.1111/j.1469-7793.1999.00645.x
复制
发表时间：
1999-11-01
影响因子：
5.5
通讯作者：
Murrell-Lagnado, RD
中科院分区：
文献类型：
--
作者：
Ho, IHM;Murrell-Lagnado, RD
1. G蛋白门控的内向纠偏K+ (GIRK)通道由G β γ和内部Na+独立激活，其机制需要磷脂酰肌醇磷酸。GIRK2中226位的天冬氨酸（Asp）对Na+依赖性激活GIRK1-GIRK2异质通道至关重要。我们在非洲爪蟾卵母细胞中表达了野生型和突变型的GIRK1-GIRK2通道，并测试了Na+和中和Asp226对通道与磷脂酰肌醇4,5-二磷酸（PIP2）功能相互作用的影响。通过将抗pip2抗体应用于内外膜斑块，GIRK2的D226N突变和增加内部[Na+]，使GIRK1-GIRK2电流的抑制率降低了10倍。2 . GIRK1 （N217D）的反向突变增加了抑制率。在20mm Na+ 0.4存在时，纯化PIP2激活的剂量-反应关系转移到较低浓度。PIP2的三种合成异构体，PI(4,5)P-2， PI(3,4)P-2和PI(3,5)P-2，以相似的能力激活了GIRK通道。我们得出结论，Na+直接与GIRK2的Asp226相互作用，降低负静电电位，促进通道与PIP2的功能相互作用。
1. G protein-gated inwardly rectifying K+ (GIRK) channels are activated independently by G beta gamma and internal Na+ via mechanisms requiring phosphatidylinositol phosphates. An aspartate (Asp) at position 226 in GIRK2 is crucial for Na+-dependent activation of GIRK1-GIRK2 heteromeric channels. We expressed wild-type and mutant GIRK1-GIRK2 channels in Xenopus oocytes and tested the effects of Na+ and neutralizing Asp226 on the functional interactions of the channels with phosphatidylinositol 4,5-bisphosphate (PIP2).2. The rate of inhibition of GIRK1-GIRK2 currents by application of anti-PIP2 antibody to inside-out membrane patches was slowed > 2-fold by the D226N mutation in GIRK2 and by increasing internal [Na+]. The reverse mutation in GIRK1 (N217D) increased the rate of inhibition.3. The dose-response relationship for activation by purified PIP2 was shifted to lower concentrations in the presence of 20 mM Na+.4. Three synthetic isoforms of PIP2, PI(4,5)P-2, PI(3,4)P-2 and PI(3,5)P-2, activated GIRK channels with similar potencies.5. We conclude that Na+ directly interacts with Asp226 of GIRK2 to reduce the negative electrostatic potential and promote the functional interaction of the channels with PIP2.